Mammalian sperm are not able to fertilize eggs immediately after ejaculation. They acquire fertilization capacity after residing in the female tract for a finite period of time. The physiological changes occuring in the female reproductive tract that render the sperm able to fertilize constitute the phenomenon of "sperm capacitation". It has been demonstrated that capacitation is associated with an increase in the tyrosine phosphorylation of a subset of proteins. It has also been demonstrated that the increase in protein tyrosine phosphorylation is regulated by a cAMP-dependent pathway involving protein kinase A (PKA). Hyperpolarization of the sperm plasma membrane has also been associated with capacitation. It is hypothesized that hyperpolarization of sperm plasma membrane during capacitation is, in some way, related to the cAMP-dependent phosphorylation of proteins. The objectives of this amended proposal are to elucidate the sequence of reactions and their mechanisms leading to hyperpolarization of mouse sperm during capacitation. The following specific aims are proposed: 1) to elucidate the role of media components in regulating sperm membrane potential and capacitation; 2) to determine the link between membrane hyperpolarization, cAMP and protein tyrosine phosphorylation; and 3) to define the regions of the sperm cell that display signal transduction during capacitation.